This CEGS will use an innovative combination of approaches to address two fundamental questions in genome biology: What do our genes do, and where did we come from? Rapid progress in genomics has provided nearly complete sequences for several organisms. Comparative analysis suggests many fundamental pathways and gene networks are conserved between organisms. And yet, the morphology, behavior, physiology, and disease susceptibility of different species are obviously and profoundly different. What are the mechanisms that generate new functions for genes, new physiological traits, and the unique form and functions of different species? Has the great variety of life forms been created by changes in gene number, by alterations in the functional attributes of particular proteins, or by diversification of the regulatory mechanisms that control where and when genes are expressed? This CEGS proposes a pioneering analysis of vertebrate diversity using a combination of techniques from structural and functional genomic and traditional genetics in zebrafish and sticklebacks. The unique experimental advantages of these two models will make it possible to take complementary approaches. The "bottom-up" approach will test the diversification in expression and genetic function of duplicated gene pairs, a major hallmark of the vertebrate genome. In situ hybridization analysis will be used to compare the expression patterns of 2500 genes. Morpholino knockout experiments will test how the functions of duplicated genes diverge, and generate a database of gene functions for many genes identified in sequencing projects. The complementary "top-down" approach will begin with naturally occurring species of sticklebacks that show profound differences in size, anatomy, and physiological traits. Genetic crosses will be used to identify the number and location of genetic changes that create the anatomical and physiological differences between recently evolved species from different regions around the world. Development of genetic and physical mapping resources for sticklebacks will make it possible to identify the actual genes and mutations responsible for evolutionary change. Immediate data release, free access to reagents, and an annual summer training course in fish genomic and genetics will ensure that the innovative approaches and results from this research will be widely disseminated to the research community. This unique combination of approaches will establish a completely new and detailed understanding of the genomic mechanisms responsible for morphological and physiological differences between living forms.